[BioC] yet another question on technical replicates...

[Giorgi, Elena]

Dear Board,

I know this topic has been discussed several times, yet I'm still
confused on how to come up with the right design matrix when technical
replicates are present, especially when dealing with affy arrays. 

For one thing, I know that when we have the same number of tech reps per
biological sample, then we can proceed and use the duplicate correlation
function, correct?

On the other hand, when this is not the case, what's the best strategy
to use? Averaging is not recommended, yet if we have 2-3 arrays per
sample, we don't have enough degrees of freedom to be able to include
the technical replication effect, isn't this so?

One example that came up in our lab was an affy experiment with two
cell-lines; for group1 we had 5 arrays, one biological replicate each,
and for group2 we had 4 arrays, 2 tech reps from one sample and 2 tech
reps from a different sample. 

We used the following design matrix:

1 0 0
1 0 0
1 0 0
1 0 0
1 0 0
0 1 0
0 1 0
0 0 1
0 0 1

And, in order to test the differences between the two groups, the
following contrast: c(-1, 0.5, 0.5).

Does this sound like a reasonable approach? In general, should we
include a different column in the design matrix for each tech rep group
and average the contrast coefficients accordingly? Or is this just
equivalent to averaging the tech reps?

Thanks so much,
Elena


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